The use of radioactively labeled diagnostic and therapeutic agents have become routine practice in clinical and analytical laboratories throughout the world. Such radioactively labeled compounds are used both in vitro, for example, in radioimmunoassay systems, and in vivo, for example, both in diagnostic imaging techniques and in radiation therapy techniques.
Initially, the number of radioisotopes that could be firmly attached to the typical organic molecules used as diagnostic and therapeutic agents was limited. The difficulty in forming stable carbon-metal bonds prevented the early utilization of many radioactive metals and typically limited radioisotopes used to label organic molecules to isotopes of phosphorus, carbon, hydrogen and iodine.
Recently, a new approach has enabled the labeling of such agents with metal ions. In this approach a chelating moiety is covalently attached to the molecule of interest, and a radioactive ion is then chelated by the sequestering groups of the chelator. Typical chelating moieties which have been used for this purpose in the prior art have been analogues or derivatives of ethylenediaminetetraacetic acid (EDTA), although many variations have also occurred.
Attempts to "mark" or "tag" cancer cells in order to differentiate them from normal tissue have been extensively investigated. Various fluorescent compounds such as tetracycline derivatives, acridine dyes and porphyrin compounds have been tried with mixed results. Of these, porphyrin compounds have shown remarkable affinity for neoplastic tissues.
Porphyrins and related analogs are complex tetrapyrrole compounds normally found in plants and in animals. They perform many vital biological functions by combining with metallic ions such as iron, magnesium, manganese, zinc, etc. to form metalloporphyrins. Metalloporphyrins are essential for the normal metabolism of plants and animals. Many of these compounds exhibit strong fluorescence when exposed to an appropriate exciting light source.
The preferential affinity of porphyrin compounds for various type of neoplasms has been known for more than four decades. When injected intravenously into tumor-bearing animals, a brilliant red-orange fluorescence is produced by ultra violet light activation of the porphyrin accumulated in the tumor.
Various porphyrin compounds have been labeled with radionuclides such as .sup.64 Cu and .sup.57 Co. Protoporphyrin and hematoporphyrin, an artificial porphyrin prepared by treating hemoglobin with concentrated sulfuric acid, labeled with .sup.64 Cu have been shown to concentrate in mouse tumors.
The common method of labeling porphyrins with radionuclides involves the reflux reaction of a porphyrin with a radioactive metallic salt in an acidic or basic medium. Dilute hydrochlorice acid, acetic acid or dilute base such as sodium hydroxide is used to dissolve the porphyrin and to act as the reaction medium. An aqueous solution of cobaltous chloride (.sup.57 CoCl.sub.2), cuprous chloride (.sup.64 CuCl.sub.2) or .sup.64 Cu-acetate is added to the porphyrin solution and refluxed for 30 minutes to up to 24-48 hours depending on the reactivity of the porphyrin used in the labeling process. The pH of the radioactive admixture is then adjusted to 6-8 whenever possible without causing denaturation or precipitation of the radiolabeled porphyrin. In many instances, the labeled product must remain in either acidic or basic condition in order to insure chemical and labeling stability.
Although the labeling process is quite simple, the labeling yield is unsatisfactory, ranging from 10-40%. The final labeled product contains many radioactive impurities. These include free or unbound radionuclide, denatured by-products and insoluble radiocolloids in the form of hydroxides such as .sup.57 Co(OH).sub.2 or .sup.64 Cu(OH).sub.2. Without extensive purification processes, these preparations are not useful or suitable for medical applications.